Incidence of Infectious Drug Resistance Among Fecal Coliforms Isolated from Raw Sewage

Raw sewage was examined for the incidence of antibiotic-resistant coliforms present among both total and fecal coliforms. In both groups, it was found that approximately 3% of the coliform bacteria were resistant to two or more antibiotics. Of these organisms, 48% were capable of transferring all or part of their antibiotic resistance to an antibiotic-sensitive, F⁻, derivative of Escherichia coli K-12. Among the R factors identified, those conferring resistance to streptomycin-tetracycline, ampicillin-streptomycin-tetracycline, and ampicillin or ampicillin-streptomycin accounted for 23, 20, and 15%, respectively, of the total R factors detected. The data indicate a significant level of infectious drug resistance among the fecal coliforms of the urban population. The data indicate further that because of the high incidence of coliform bacteria found to be doubly resistant to streptomycin and tetracyline, the inclusion of these antibiotics in selective media used for routine total or fecal coliform counts may serve to identify domestic sources of pollution.     

Evaluation of Standard and Modified M-FC, MacConkey, and Teepol Media for Membrane Filtration Counting of Fecal Coliforms in Water

MacConkey agar, standard M-FC agar, M-FC agar without rosolic acid, M-FC agar with a resuscitation top layer, Teepol agar, and pads saturated with Teepol broth, were evaluated as growth media for membrane filtration counting of fecal coliform bacteria in water. In comparative tests on 312 samples of water from a wide variety of sources, including chlorinated effluents, M-FC agar without rosolic acid proved the medium of choice because it generally yielded the highest counts, was readily obtainable, easy to prepare and handle, and yielded clearly recognizable fecal coliform colonies. Identification of 1,139 fecal coliform isolates showed that fecal coliform tests cannot be used to enumerate Escherichia coli because the incidence of E. coli among fecal coliforms varied from an average of 51% for river water to 93% for an activated sludge effluent after chlorination. The incidence of Klebsiella pneumoniae among fecal coliforms varied from an average of 4% for the activated sludge effluent after chlorination to 32% for the river water. The advantages of a standard membrane filtration procedure for routine counting of fecal coliforms in water using M-FC agar without rosolic acid as growth medium, in the absence of preincubation or resuscitation steps, are outlined.     

Antimicrobial resistant coliform bacteria in the Gomti river water and determination of their tolerance level

The distribution of resistance to ampicillin, chloramphenicol, sulfonamides, tetracycline, and streptomycin among coliform in the Gomti river water samples was investigated. The coliform populations were isolated on Mac Conky and eosin methylene blue (EMB) agar plates supplemented with antibiotics. The incidence of resistance among the coliform population varied considerably in different drug and water sampling sites. Coliform bacteria showed lower drug resistant viable count in sampling site-III (receiving treated wastewater) as compared to more polluted site-I and site-II. Viable count of coliform population obtained on both medium was recorded higher against erythromycin from sampling site-III. Lower viable count of coliforms was recorded against tetracycline in site-II and III. Similar resistance pattern was obtained in the frequency of E. coli and Enterobacter species from all the three sampling sites. Percentage of antibiotic resistant E. coli was observed higher than Enterobacter spp among the total coliforms against all antibiotics tested without Erythromycin and penicillin in site-I and II respectively. Isolates of E. coli and Enterobacter spp. showed their tolerance level (MIC) in the range of 2-100 against the antibiotics tested. Maximum number of isolates of both genus exhibited their MICs at lower concentration range 2-5µg/ml against ciprofloxacin, tetracyclin and amoxycillin.

Abbreviations

EMB - Eosin methylene blue, IMViC tests - Indole, Methyl Red, Voges Proskauer and Citrate Utilization Tests, MIC - Minimum inhibitory concentration.     

Incidence of coliphage in potable water supplies.

Samples of drinking water from different sources in greater Cairo, Egypt, and bottled drinking water were tested for total coliform, fecal coliform, and coliphage populations. Of the 147 samples tested, 4 samples were positive for both total coliforms and coliphage, 65 samples were negative for total coliforms, fecal coliforms, and coliphage, and 78 samples were positive for coliphage and negative for total coliforms and fecal coliforms. The incidence of coliphage in these potable water supplies reflects the probability of human pathogenic virus survival in these waters also.     

Incidence of coliphage in potable water supplies

Samples of drinking water from different sources in greater Cairo, Egypt, and bottled drinking water were tested for total coliform, fecal coliform, and coliphage populations. Of the 147 samples tested, 4 samples were positive for both total coliforms and coliphage, 65 samples were negative for total coliforms, fecal coliforms, and coliphage, and 78 samples were positive for coliphage and negative for total coliforms and fecal coliforms. The incidence of coliphage in these potable water supplies reflects the probability of human pathogenic virus survival in these waters also.     

Classification of antibiotic resistance patterns of indicator bacteria by discriminant analysis: use in predicting the source of fecal contamination in subtropical waters

The antibiotic resistance patterns of fecal streptococci and fecal coliforms isolated from domestic wastewater and animal feces were determined using a battery of antibiotics (amoxicillin, ampicillin, cephalothin, chlortetracycline, oxytetracycline, tetracycline, erythromycin, streptomycin, and vancomycin) at four concentrations each. The sources of animal feces included wild birds, cattle, chickens, dogs, pigs, and raccoons. Antibiotic resistance patterns of fecal streptococci and fecal coliforms from known sources were grouped into two separate databases, and discriminant analysis of these patterns was used to establish the relationship between the antibiotic resistance patterns and the bacterial source. The fecal streptococcus and fecal coliform databases classified isolates from known sources with similar accuracies. The average rate of correct classification for the fecal streptococcus database was 62.3%, and that for the fecal coliform database was 63.9%. The sources of fecal streptococci and fecal coliforms isolated from surface waters were identified by discriminant analysis of their antibiotic resistance patterns. Both databases identified the source of indicator bacteria isolated from surface waters directly impacted by septic tank discharges as human. At sample sites selected for relatively low anthropogenic impact, the dominant sources of indicator bacteria were identified as various animals. The antibiotic resistance analysis technique promises to be a useful tool in assessing sources of fecal contamination in subtropical waters, such as those in Florida.     

Antibiotic resistance patterns of fecal coliforms isolated from domestic sewage before and after treatment in an aerobic lagoon.

The resistance of 260 strains of fecal coliforms, isolated from raw domestic sewage and aerobic lagoon effluent, to ampicillin, aureomycin, chloromycetin, gentamicin, streptomycin, sulfadiazine, and tetracycline, was determined. Aerobic lagoon treatment produced a 20-fold reduction in the fecal coliform numbers. No statistically significant difference in antibiotic resistance was observed between the fecal coliforms found in raw sewage and in lagoon effluent despite a trend towards the loss of resistance in the latter. Antibiotic resistance, either single or multiple, did not contribute to, or detract from, bacterial survival in the aerobic lagoon. Of the isolates, 15% showed resistance to two or more antiobiotics.     

A note on antibiotic resistance in the bacterial flora of raw sewage and sewage-polluted River Tigris in Mosul, Iraq

Polluted water samples collected from the River Tigris in the vicinity of a raw sewage outfall were examined for the incidence of antibiotic resistance among coliform bacteria on three occasions during 1983. Eighty percent or more of the coliform bacteria were resistant to one or more antibiotics. At the same time, raw sewage samples were examined for the incidence of antibiotic-resistant bacteria, and Escherichia coli, Pseudomonas spp. and Staphylococcus spp. were selected for sensitivity testing. Collectively, more than 90% of the 480 strains of the three organisms were resistant to one or more antibiotics. The minimal inhibitory concentration (MIC) of ampicillin for twenty-nine strains including coliforms, E. coli, Klebsiella sp., Serratia sp., Ps. aeruginosa, Pseudomonas sp., Micrococcus sp., Staph. aureus, Streptococcus faecalis and Bacillus sp. from raw sewage and polluted River Tigris water was determined and that for Ps. aeruginosa was 250 micrograms/ml. The high incidence of antibiotic-resistant bacteria in natural waters could be related to the widespread use of antibiotics in this locality.     

A note on antibiotic resistance in the bacterial flora of raw sewage and sewage-polluted River Tigris in Mosul, Iraq

Polluted water samples collected from the River Tigris in the vicinity of a raw sewage outfall were examined for the incidence of antibiotic resistance among coli-form bacteria on three occasions during 1983. Eighty percent or more of the coli-form bacteria were resistant to one or more antibiotics. At the same time, raw sewage samples were examined for the incidence of antibiotic-resistant bacteria, and Escherchia coli, Pseudomonas spp. and Staphylococcus spp. were selected for sensitivity testing. Collectively, more than 90% of the 480 strains of the three organisms were resistant to one or more antibiotics. The minimal inhibitory concentration (MIC) of ampicillin for twenty-nine strains including coliforms, E. coli, Klebsiella sp., Serratia sp., Ps. aeruginosa, Pseudomonas sp., Micrococcus sp., Staph. aureus, Streptococcus faecalis and Bacillus sp. from raw sewage and polluted River Tigris water was determined and that for Ps. aeruginosa was 250 μg/ml. The high incidence of antibiotic-resistant bacteria in natural waters could be related to the widespread use of antibiotics in this locality.     

Enumeration of total coliforms and Escherichia coli from source water by the defined substrate technology

Many water utilities are required to monitor source water for the presence of total coliforms, fecal coliforms, or both. The Colilert system, an application of the defined substrate technology, simultaneously detects the presence of both total coliforms and Escherichia coli directly from a water sample. After incubation, the formula becomes yellow if total coliforms are present and fluorescent at 366 nm if E. coli is in the same sample. No confirmatory tests are required. The Colilert system was previously assessed with distribution water in a national evaluation in both most-probably-number and presence-absence formats and found to produce data equivalent to those obtained by using Standard Methods for the Examination of Water and Wastewater (Standard Methods). The Colilert system was now compared with Standard Methods multiple-tube fermentation (MTF) for the enumeration of total coliforms and E. coli from surface water. All MTF tubes were confirmed according to Standard Methods, and subcultures were made to identify isolates to the species level. The Colilert system was found equally sensitive to MTF testing by regression, t test, chi-square, and likelihood fraction analyses. Specificity of the Colilert system was shown by the isolation of a species of total coliform or E. coli after the appropriate color change. The Colilert test can be used for source water samples when enumeration is required, and the benefits previously described for distribution water testing--sensitivity, specificity, less labor, lower cost, faster results, no noncoliform heterotroph interference--are applicable to this type of water analysis.     

Enumeration of total coliforms and Escherichia coli from source water by the defined substrate technology.

Many water utilities are required to monitor source water for the presence of total coliforms, fecal coliforms, or both. The Colilert system, an application of the defined substrate technology, simultaneously detects the presence of both total coliforms and Escherichia coli directly from a water sample. After incubation, the formula becomes yellow if total coliforms are present and fluorescent at 366 nm if E. coli is in the same sample. No confirmatory tests are required. The Colilert system was previously assessed with distribution water in a national evaluation in both most-probably-number and presence-absence formats and found to produce data equivalent to those obtained by using Standard Methods for the Examination of Water and Wastewater (Standard Methods). The Colilert system was now compared with Standard Methods multiple-tube fermentation (MTF) for the enumeration of total coliforms and E. coli from surface water. All MTF tubes were confirmed according to Standard Methods, and subcultures were made to identify isolates to the species level. The Colilert system was found equally sensitive to MTF testing by regression, t test, chi-square, and likelihood fraction analyses. Specificity of the Colilert system was shown by the isolation of a species of total coliform or E. coli after the appropriate color change. The Colilert test can be used for source water samples when enumeration is required, and the benefits previously described for distribution water testing--sensitivity, specificity, less labor, lower cost, faster results, no noncoliform heterotroph interference--are applicable to this type of water analysis.     

Frequency of virulence genes and antibiotic resistances in Enterococcus spp. isolates from wastewater and feces of domesticated mammals and birds, and wildlife

Enterococci are gastrointestinal tract residents and also an important cause of nosocomial infections. To understand which species, virulence determinants, and antibiotic resistances are prevalent in enterococci shed by various hosts groups, a total of 1460 strains isolated from 144 fecal samples obtained from wastewater, domesticated mammals and birds, and wildlife were characterized. Identification of isolates to the species level showed that Enterococcus faecalis was dominant in domesticated mammals and birds and wildlife feces, whereas Enterococcus faecium was dominant among wastewater isolates, and that no single Enterococcus species could be associated with a specific host group. The frequency of 12 virulence determinants was evaluated among isolates, but no single virulence determinant could be associated with a specific host group. Resistance to 12 antibiotics was evaluated among isolates, and it was shown that the highest frequency of resistance at breakpoint concentration was found in domesticated mammals and birds (P?≤ 0.05 for 4 antibiotics). Our results suggests that (1) species identification and virulence typing of Enterococcus spp. isolates are not useful for the identification of the host groups responsible for fecal contamination of water by microbial source tracking and that (2) antibiotic use for clinical, veterinary, or animal husbandry practices is promoting resistance.     

Effect of dredge spoil deposition on fecal coliform counts in sediments at a disposal site.

The most-probable-number of fecal coliforms in sediments was monitored at the New London dump site in Long Island Sound during the deposition of dredge spoil from the Thames River. Although the geometric mean for fecal coliforms at five stations in the river was 14,000/100 ml before dredging commenced, the deposition of this material did not increase the incidence of fecal coliforms at 17 spoil stations and 13 control stations in the disposal and surrounding areas. Fecal coliforms appear to occur only in the surface sediment material and are diluted by the subsurface material during the dredging operation. Fecal coliform analyses of bottom waters during high and low tides indicated that the flow of water from the Thames River played a major role in determining the most-probable-number of fecal coliforms in the sediments at the disposal site.     

Effect of dredge spoil deposition on fecal coliform counts in sediments at a disposal site.

The most-probable-number of fecal coliforms in sediments was monitored at the New London dump site in Long Island Sound during the deposition of dredge spoil from the Thames River. Although the geometric mean for fecal coliforms at five stations in the river was 14,000/100 ml before dredging commenced, the deposition of this material did not increase the incidence of fecal coliforms at 17 spoil stations and 13 control stations in the disposal and surrounding areas. Fecal coliforms appear to occur only in the surface sediment material and are diluted by the subsurface material during the dredging operation. Fecal coliform analyses of bottom waters during high and low tides indicated that the flow of water from the Thames River played a major role in determining the most-probable-number of fecal coliforms in the sediments at the disposal site.     

Longitudinal study of microbial diversity and seasonality in the Mexico City metropolitan area water supply system

In the Mexico City metropolitan area (MCMA), 70% of the water for 18 million inhabitants is derived from the Basin of Mexico regional aquifer. To provide an overview of the quality of the groundwater, a longitudinal study was conducted, in which 30 sites were randomly selected from 1,575 registered extraction wells. Samples were taken before and after chlorine disinfection during both the rainy and dry seasons (2000-2001). Microbiological parameters (total coliforms, fecal coliforms, streptococci, and Vibrio spp.), the presence of Helicobacter pylori, and physicochemical parameters, including the amount of trihalomethanes (THMs), were determined. Although microorganisms and inorganic and organic compounds were evident, they did not exceed current permissible limits. Chlorine levels were low, and the bacterial counts were not affected by chlorine disinfection. Eighty-four bacterial species from nine genera normally associated with fecal contamination were identified in water samples. H. pylori was detected in at least 10% of the studied samples. About 40% of the samples surpassed the THM concentration allowed by Mexican and U.S. regulations, with levels of chloroform being high. The quality of the water distributed to the MCMA varied between the rainy and dry seasons, with higher levels of pH, nitrates, chloroform, bromodichloromethane, total organic carbon, and fecal streptococci during the dry season. This study showed that the groundwater distribution system is susceptible to contamination and that there is a need for a strict, year-round disinfection strategy to ensure adequate drinking-water quality. This situation in one of the world's megacities may reflect what is happening in large urban centers in developing countries which rely on a groundwater supply.     

Antibiotic resistance among coliforms and Pseudomonas spp. from bodies of water around Port Harcourt, Nigeria

Samples from municipal waste water, the Bonny River estuary and wells in and around Port Harcourt were examined for bacteriological quality over a 9 month period. A total of 157 Pseudomonas spp., 133 Escherichia coli and 282 other coliforms were isolated and tested for the incidence of resistance to 10 antibiotics. All of the Pseudomonas spp. were resistant to at least one of the antibiotics while 96.2% were resistant to two or more. Most (83.5%) of the E. coli and other coliforms (91.8%) were resistant to at least one antibiotic. All strains were susceptible to gentamicin. Minimal inhibitory concentrations of ampicillin and tetracycline for E. coli ranged from 6.25 to 50 and 6.25 to 12.5 micrograms/ml, respectively. Minimal inhibitory concentrations of ampicillin and tetracycline were 1000 and 25 micrograms/ml for the Pseudomonas strains. The high incidence of bacterial resistance to antibiotics is discussed in relation to the widespread use of antibiotics, and possible public health implications.     

Bacterial pollution indicators in the intestinal tract of freshwater fish

A study was made of the occurrence, distribution, and persistence of coliforms, fecal coliforms, and fecal streptococci in the intestinal tract of freshwater fish. A total of 132 fish representing 14 different species were used in various phases of these experiments. Examination of the intestinal contents of 78 fish from moderately polluted sections of the Little Miami River indicated that fecal coliform densities were lowest in bluegills (less than 20 per gram) and highest in catfish (1,090,000 per gram). Levels of fecal streptococci for these two species were 220 and 240,000 per gram, respectively. The occurrence of fecal coliforms in fish caught in this stream reflected the warm-blooded-animal-pollution level of the water. All fish used in this phase of the study were caught during July, August, and September when the water temperatures were between 13 and 18 C. The fate of fecal coliforms and Streptococcus faecalis in the fish intestine indicated that these organisms can probably survive and multiply when fish and water temperatures are 20 C or higher, but only when the organisms are retained in the gut for periods beyond 24 hr. Based on the biochemical reactions for 3,877 coliform strains isolated from 132 freshwater fish of 14 different species, 91.4% of all strains were composed of five IMViC types. In a similar study of the biochemical reactions of 850 streptococci isolated from the intestinal tract of 55 freshwater fish, the predominant strains included S. faecalis and various closely associated biotypes. No consistently recurring pattern for either coliforms or streptococci could be developed to identify species of fish investigated. The composition of the intestinal flora is, however, related in varying degree to the level of contamination of water and food in the environment.     

Antibiotic resistant bacteria in Windermere and two remote upland tarns in the English Lake District

The incidence of antibiotic resistance was determined in over 2000 bacteria which were divided into the following groups: faecal streptococci, coliforms (excluding Escherichia coli), E. coli, Pseudomonas spp. and aquatic bacteria (i.e. bacteria predominant in the lake water which were excluded from the previous four categories). The isolates were obtained from the water of Windermere (English Lake District) and from a sewage effluent which entered the lake. With the exception of the faecal streptococci, the incidence of antibiotic resistance was higher in the bacteria isolated from the lake water than in those from the effluent, and ranked according to groups Pseudomonas spp. greater than E. coli greater than aquatic bacteria greater than coliforms greater than faecal streptococci. The highest incidence of multiple resistance was found among the pseudomonads. When corrected for the relative size of each population the pool of antibiotic resistance in the aquatic bacteria was by far the largest. The incidence of antibiotic resistance in aquatic bacteria isolated from Windermere was, however, lower than in those isolated from two remote upland tarns. This finding may have been due to differences in the species composition of the three sites except that the same results were obtained when only fluorescent pseudomonads were tested. The upland tarns were not totally isolated from man and other animals but did not receive any sewage or other effluents and therefore the results were surprising. Possible explanations include a lack of susceptibility in aquatic bacteria and increased resistance associated with growth in nutrient poor environments.     

R factors in coliform-fecal coliform sewage flora of the prairies and Northwest Territories of Canada.

Coliform and fecal coliform populations found in the raw sewages and final sewage effluents of the prairie provinces and the Northwest Territories were examined for antibiotic resistance and the possession of R factors. It was determined that 8.91% of the total coliform and 10.80% of the fecal coliform populations carried R factors. The following numbers of combinations of R determinants were found: 39 in the Escherichia coli population, 6 in the Citrobacter population, 20 in the Enterobacter populations, 10 in the Klebsiella populations, and 11 in the Aeromonas populations. The maximum number of R determinants transferable simultaneously was seven; organisms with R factors containing determinants for chloramphenicol usually contained determinants for ampicillin. Of the coliform and fecal coliform populations, 2 to 4% were resistant to chloramphenicol in some provinces, and from 17 to 30% of the populations were resistant to three or more antibiotics. It was calculated that coliforms containing R factors in the raw sewage reached population levels of 1.5 X 10(7)/100 ml, and fecal coliforms containing R factors reached population levels of 8.6 X 10(5) ml. Final effluent discharges to the receiving environment contained R factor-containing coliform and fecal coliform populations of 3.1 X 10(4)/100 ml and 5.8 X 10(2)/100 ml, respectively. The incidence of bacteria containing R factors in sewage appears to be increasing with time, and their removal from sewage before discharge to the receiving environment is desirable. Consideration of data on bacteria with R factors should be made in future water quality deliberations and in discharge regulations.     

Virus occurrence in municipal groundwater sources in Quebec, Canada

A 1 year study was undertaken on groundwater that was a source of drinking water in the province of Quebec, Canada. Twelve municipal wells (raw water) were sampled monthly during a 1 year period, for a total of 160 samples. Using historic data, the 12 sites were categorized into 3 groups: group A (no known contamination), group B (sporadically contaminated by total coliforms), and group C (historic and continuous contamination by total coliforms and (or) fecal coliforms). Bacterial indicators (total coliform, Escherichia coli, enteroccoci), viral indicators (somatic and male-specific coliphages), total culturable human enteric viruses, and noroviruses were analyzed at every sampling site. Total coliforms were the best indicator of microbial degradation, and coliform bacteria were always present at the same time as human enteric viruses. Two samples contained human enteric viruses but no fecal pollution indicators (E. coli, enterococci, or coliphages), suggesting the limited value of these microorganisms in predicting the presence of human enteric viruses in groundwater. Our results underline the value of historic data in assessing the vulnerability of a well on the basis of raw water quality and in detecting degradation of the source. This project allowed us to characterize the microbiologic and virologic quality of groundwater used as municipal drinking water sources in Quebec.